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The visualization of green fluorescent protein (GFP) fusions with microtubule or actin filament (F-actin) binding proteins has provided new insights into the function of the cytoskeleton during plant development. For studies on actin, GFP fusions to talin have been the most generally used reporters. Although GFP-Talin has allowed in vivo F-actin imaging in a variety of plant cells, its utility in monitoring F-actin in stably transformed plants is limited particularly in developing roots where interesting actin dependent cell processes are occurring. In this study, we created a variety of GFP fusions to Arabidopsis Fimbrin 1 (AtFim1) to explore their utility for in vivo F-actin imaging in root cells and to better understand the actin binding properties of AtFim1 in living plant cells. Translational fusions of GFP to full-length AtFim1 or to some truncated variants of AtFim1 showed filamentous labeling in transient expression assays. One truncated fimbrin-GFP fusion was capable of labeling distinct filaments in stably transformed Arabidopsis roots. The filaments decorated by this construct were highly dynamic in growing root hairs and elongating root cells and were sensitive to actin disrupting drugs. Therefore, the fimbrin-GFP reporters we describe in this study provide additional tools for studying the actin cytoskeleton during root cell development. Moreover, the localization of AtFim1-GFP offers insights into the regulation of actin organization in developing roots by this class of actin cross-linking proteins.

The actin cytoskeleton has been proposed to be a major player in plant gravitropism. However, understanding the role of actin in this process is far from complete. To address this problem, we conducted an analysis of the effect of Latrunculin B (Lat B), a potent actin-disrupting drug, on root gravitropism using various parameters that included detailed curvature kinetics, estimation of gravitropic sensitivity, and monitoring of curvature development after extended clinorotation. Lat B treatment resulted in a promotion of root curvature after a 90°reorientation in three plant species tested. More significantly, the sensitivity of maize (Zea mays) roots to gravity was enhanced after actin disruption, as determined from a comparison of presentation time of Lat B-treated versus untreated roots. A short 10-min gravistimulus followed by extended rotation on a 1-rpm clinostat resulted in extensive gravitropic responses, manifested as curvature that often exceeded 90°. Application of Lat B to the cap or elongation zone of maize roots resulted in the disruption of the actin cytoskeleton, which was confined to the area of localized Lat B application. Only roots with Lat B applied to the cap displayed the strong curvature responses after extended clinorotation. Our study demonstrates that disrupting the actin cytoskeleton in the cap leads to the persistence of a signal established by a previous gravistimulus. Therefore, actin could function in root gravitropism by providing a mechanism to regulate the proliferation of a gravitropic signal originating from the cap to allow the root to attain its correct orientation or set point angle.Plants respond to an array of environmental and developmental stimuli with gravity being one of the more significant cues to which plants must adapt to survive. Under the Earth's gravitational field, shoots normally grow upward to maximize light absorption for photosynthesis, whereas roots grow down for optimal water and nutrient acquisition. This directional growth response of plant organs to gravity or gravitropism has been conveniently divided into a series of events consisting of gravity sensing, signal transduction, signal transmission, and the growth response (Kiss, 2000). In roots, gravity sensing is believed to occur in the root cap (Sack, 1997), but recent evidence suggests that roots may have dual gravity sensors, one of which may be located in a region outside the cap (Wolverton et al., 2002a(Wolverton et al., , 2002b.Despite recent proposals of alternative gravitysensing sites in roots (Wolverton et al., 2002a), there is a great deal of cell biological and physiological evidence demonstrating that starch-containing amyloplasts in the columella region of the root cap are significant for gravity sensing (for review, see Kiss, 2000;Boonsirichai et al., 2002). Popularly known as the starch-statolith hypothesis, the sedimentation of amyloplasts within the columella cells is proposed to constitute one of the initial acts of gravity sensing in roots (Sack, 1997;Kiss, 2000). In shoots, sedi...

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